Summary of Work: The highly conserved, ubiquitous DNA repair pathway known as base excision repair(BER) is one of the key cellular defense mechanisms against deleterious effects of metabolism and environmental exposures. For example, mutations in E. coli and mammalian BER genes alike cause cells to suffer genomic instability upon environmental exposure. Among the most important sources of genomic damage repaired by BER are: hydrolytic loss of DNA bases; oxidation of bases and sugars; alkylation of DNA bases and DNA phosphate groups; and misincorporation errors during DNA replication. Generally, BER repaired lesions are smaller lesions in DNA bases, instead of large, bulky substitutions at bases, strand breaks or inter-strand cross-links. Our hypothesis is that lack of appropriate BER in mammalian cells can play a critical role in carcinogenesis and other degenerative conditions. We have recapitulated the main mammalian BER pathway in vitro using the four purified human proteins, uracil-DNA glycosylase, AP endonuclease, DNA polymerase - and DNA ligase I. We have cloned the human and mouse genes for these four enzymes, expressed the recombinant human proteins in E. coli and insect cells, and prepared cell lines with genetic alterations in each gene. Transgenic mouse models are being prepared for study of the cellular and tissue requirements for each enzyme. This project includes studies of the cellular role of BER in DNA repair, apoptosis, mutagenesis, chromosome stability, and cell signalling, growth control, human and mouse carcinogenesis.